Impact of cancer-associated mutations in CC chemokine receptor 2 on receptor function and antagonism.

CC chemokine receptor 2 (CCR2), a G protein-coupled receptor, plays a role in many cancer-related processes such as metastasis formation and immunosuppression. Since âˆ¼ 20 % of human cancers contain mutations in G protein-coupled receptors, ten cancer-associated CCR2 mutants obtained from the Genome Data Commons were investigated for their effect on receptor functionality and antagonist binding. Mutations were selected based on either their vicinity to CCR2's orthosteric or allosteric binding sites or their presence in conserved amino acid motifs. One of the mutant receptors, namely S101P2.63 with a mutation near the orthosteric binding site, did not express on the cell surface. All other studied mutants showed a decrease in or a lack of G protein activation in response to the main endogenous CCR2 ligand CCL2, but no change in potency was observed. Furthermore, INCB3344 and LUF7482 were chosen as representative orthosteric and allosteric antagonists, respectively. No change in potency was observed in a functional assay, but mutations located at F1163.28 impacted orthosteric antagonist binding significantly, while allosteric antagonist binding was abolished for L134Q3.46 and D137N3.49 mutants. As CC chemokine receptor 2 is an attractive drug target in cancer, the negative effect of these mutations on receptor functionality and drugability should be considered in the drug discovery process.

Human skin barrier formation takes place via a cubic to lamellar lipid phase transition as analyzed by cryo-electron microscopy and EM-simulation.

The skin's permeability barrier consists of stacked lipid sheets of splayed ceramides, cholesterol and free fatty acids, positioned intercellularly in the stratum corneum. We report here on the early stage of skin barrier formation taking place inside the tubuloreticular system in the secretory cells of the topmost viable epidermis and in the intercellular space between viable epidermis and stratum corneum. The barrier formation process was analysed in situ in its near-native state, using cryo-EM combined with molecular dynamics modeling and EM simulation. Stacks of lamellae appear towards the periphery of the tubuloreticular system and they are closely associated with granular regions. Only models based on a bicontinuous cubic phase organization proved compatible with the granular cryo-EM patterns. Only models based on a dehydrated lamellar phase organization agreed with the lamellar cryo-EM patterns. The data support that human skin barrier formation takes place via a cubic to lamellar lipid phase transition.

A randomized, phase 2 study of cetuximab plus cisplatin with or without paclitaxel for the first-line treatment of patients with recurrent and/or metastatic squamous cell carcinoma of the head and neck.

BACKGROUND: B490 (EudraCT# 2011-002564-24) is a randomized, phase 2b, noninferiority study investigating the efficacy and safety of first-line cetuximab plus cisplatin with/without paclitaxel (CetCis versus CetCisPac) in patients with recurrent and/or metastatic squamous cell carcinoma of the head and neck (R/M SCCHN). PATIENTS AND METHODS: Eligible patients had confirmed R/M SCCHN (oral cavity/oropharynx/larynx/hypopharynx/paranasal sinus) and no prior therapy for R/M disease. Cetuximab was administered on day 1 (2-h infusion, 400 mg/m2), then weekly (1-h infusions, 250 mg/m2). Cisplatin was given as a 1-h infusion (CetCis arm: 100 mg/m2; CetCisPac arm: 75 mg/m2) on day 1 of each cycle for a maximum of six cycles. Paclitaxel was administered as a 3-h infusion (175 mg/m2) on day 1 of each cycle. After six cycles, maintenance cetuximab was administered until disease progression or unacceptable toxicity. The primary end point was progression-free survival (PFS). We assumed a noninferiority margin of 1.40 as compatible with efficacy. RESULTS: A total of 201 patients were randomized 1 : 1 to each regimen; 191 were assessable. PFS with CetCis (median, 6 months) was noninferior to PFS with CetCisPac (median, 7 months) [HR for CetCis versus CetCisPac 0.99; 95% CI: 0.72-1.36, P = 0.906; margin of noninferiority (90% CI of 1.4) not reached]. Median overall survival was 13 versus 11 months (HR = 0.77; 95% CI: 0.53-1.11, P = 0.117). The overall response rates were 41.8% versus 51.7%, respectively (OR = 0.69; 95% CI: 0.38-1.20, P = 0.181). Grade ≥3 adverse event rates were 76% and 73% for CetCis versus CetCisPac, respectively, while grade 4 toxicities were lower in the two-drug versus three-drug arm (14% versus 33%, P = 0.015). No toxic death or sepsis were reported and cardiac events were negligible (1%). CONCLUSION: The two-drug CetCis regimen proved to be noninferior in PFS to a three-drug combination with CetCisPac. The median OS of both regimens is comparable with that observed in EXTREME, while the life-threatening toxicity rate appeared reduced. CLINICAL TRIAL NUMBER: EudraCT# 2011-002564-24.

Texture of cellulose microfibrils of root hair cell walls of Arabidopsis thaliana, Medicago truncatula, and Vicia sativa.

Cellulose is the most abundant biopolymer on earth, and has qualities that make it suitable for biofuel. There are new tools for the visualisation of the cellulose synthase complexes in living cells, but those do not show their product, the cellulose microfibrils (CMFs). In this study we report the characteristics of cell wall textures, i.e. the architectures of the CMFs in the wall, of root hairs of Arabidopsis thaliana, Medicago truncatula and Vicia sativa and compare the different techniques we used to study them. Root hairs of these species have a random primary cell wall deposited at the root hair tip, which covers the outside of the growing and fully grown hair. The secondary wall starts between 10 (Arabidopsis) and 40 (Vicia) µm from the hair tip and the CMFs make a small angle, Z as well as S direction, with the long axis of the root hair. CMFs are 3-4 nm wide in thin sections, indicating that single cellulose synthase complexes make them. Thin sections after extraction of cell wall matrix, leaving only the CMFs, reveal the type of wall texture and the orientation and width of CMFs, but CMF density within a lamella cannot be quantified, and CMF length is always underestimated by this technique. Field emission scanning electron microscopy and surface preparations for transmission electron microscopy reveal the type of wall texture and the orientation of individual CMFs. Only when the orientation of CMFs in subsequent deposited lamellae is different, their density per lamella can be determined. It is impossible to measure CMF length with any of the EM techniques.

Sleep quality, estradiol levels, and behavioral factors in late reproductive age women.

OBJECTIVE: To estimate the prevalence of perceived poor sleep in women aged 35-49 years and to correlate sleep quality with levels of gonadal steroids and predictors of poor sleep. METHODS: A cohort of 218 black and 218 white women aged 35-47 years at enrollment (aged 37-49 at final follow-up) with regular menstrual cycles was identified through random digit dialing for a longitudinal study of ovarian aging correlates. Data obtained at four assessment periods, including enrollment, over a 2-year interval were collected between days 1 and 6 (mean = 3.9) of the menstrual cycle. The primary outcome measure was subjects' rating of sleep quality at each assessment period. Associations of sleep quality with hormone levels (estradiol, follicle-stimulating hormone, luteinizing hormone, testosterone, and dehydroepiandrosterone sulfate) and other clinical, behavioral, and demographic variables were examined in bivariable and multivariable analyses. RESULTS: Approximately 17% of subjects reported poor sleep at each assessment period. Significant independent associations with poor sleep included greater incidence of hot flashes (odds ratio [OR] 1.52; 95% confidence interval [CI] 1.08, 2.12, P =.02), higher anxiety levels (OR 1.03; 95% CI 1.00, 1.06, P =.04), higher depression levels (OR 1.05; 95% CI 1.02, 1.07, P <.001), greater caffeine consumption (OR 1.25; 95% CI 1.04, 1.49, P =.02), and lower estradiol levels in women aged 45-49 (OR 0.53; 95% CI 0.34, 0.84, P =.006), after adjustment for current use of sleep medications. CONCLUSION: Both hormonal and behavioral factors were associated with sleep quality. Estradiol levels are an important factor in poor sleep reported by women in the 45-49 age group. Further evaluation of estrogen treatment for poor sleep of women 45 years and older is warranted.

Hot flashes in the late reproductive years: risk factors for Africa American and Caucasian women.

Hot flashes are a primary reason that midlife women seek medical care, but there is little information about the onset or the predictors of hot flashes in the years before the menopause. This study examines women's experience of hot flashes in the late reproductive years, comparing African American and Caucasian women, and identifies hormonal, behavioral, and environmental risk factors for hot flashes associated with ovarian aging. Data are from a population-based prospective cohort study of ovarian aging in women who were ages 35--47, in general good health, and had regular menstrual cycles at study enrollment. Hot flashes were assessed by subject report in a structured interview at the first follow-up period and correlated highly with previous prospective daily ratings of hot flashes (p = 0.0001). Blood samples were obtained in the first 6 days of the menstrual cycle in two consecutive cycles at enrollment and two consecutive cycles at follow-up. Predictor variables include hormone measures, structured interview, and standard questionnaire data. Thirty-one percent of the sample (n = 375) reported hot flashes (mean age 41 years). In bivariate analysis, more African American than Caucasian women reported hot flashes (38% vs. 25%, p = 0.01). Significant predictors of hot flashes in the final multivariable logistic regression model were higher follicle-stimulating hormone (FSH) levels (odds ratio [OR] 3.19), anxiety (OR 1.06), baseline menopausal symptoms (OR 4.91), alcohol use (OR 1.09), body mass index (BMI) (OR 1.04), and parity (OR 1.20). Race did not predict hot flashes after adjusting for these variables. Hot flashes commonly occur before observable menstrual irregularities in the perimenopause and are associated with both hormonal and behavioral factors. The association of hot flashes with increased body mass (BMI) challenges the current "thin" hypothesis and raises important questions about the role of BMI in hormone dynamics in the late reproductive years.

Symptom reports from a cohort of African American and white women in the late reproductive years.

OBJECTIVE: To identify symptoms experienced in a cohort of healthy women in the late reproductive years; to compare symptom reports between African American and Caucasian women; and to determine the extent to which other factors in reproductive health, mood and behavior, lifestyle, and demographic background are associated with the reported symptoms. DESIGN: A cohort of women aged 35 to 47 years (mean age, 41 years) was identified through random digit dialing. This study is a cross-sectional analysis of data collected at enrollment from a subset of 308 women who completed daily symptom reports (DSR) for one menstrual cycle. Data were obtained in structured interviews and self-administered standard questionnaires. The associations of the study variables with symptoms as assessed by the DSR were examined using analysis of variance and general linear models. RESULTS: The African American women were significantly more likely to report in interview that they experienced menopausal symptoms (46% vs. 30%; p < 0.001) and had significantly higher ratings on the physiological symptom factor of the DSR, which included hot flashes, dizziness, poor coordination/clumsiness, urine leaks, and vaginal dryness. The DSR yielded two other factors of psychological and somatic symptoms. Race was associated only with the physiological symptom factor in the multivariable analyses. Neither race nor age were associated with psychological symptoms, which were predicted by current or past mood problems. CONCLUSIONS: Symptoms commonly associated with the menopause are experienced in the late reproductive years before observable changes in menstrual cycles. African American women reported more physiological symptoms than white women. These data provide an essential baseline for longitudinal study of symptoms associated with the ovarian decline in the perimenopausal years.

Epidemiological data of Crohn's disease in Israel: etiological implications.

We have reviewed the public health aspects of Crohn's disease in Israel. Crohn's disease has increased in frequency in the Jewish population of Israel over the past twenty years, although it is still less common than ulcerative colitis. The increase has occurred in both sexes and in European- and American-born, Asian- and African-born, and Israeli-born Jews. Crohn's disease is, however, less prevalent in Israel than in Jews living abroad, with the exception of European- and American-born Jews who have a high frequency of the disease. Jews originating from middle European countries are at a particularly high risk. The peak age at onset is 20-29 years, with Israeli-born persons developing the disease at a relatively early age. Intestinal cancer is a rare complication in this country. The rate of familial involvement is apparently lower than that reported elsewhere. There are no adequate data on environmental factors causing the disease. The disease remains rare in the Arab population.

Comparison of an enzyme-linked immunosorbent assay using monoclonal antibodies and a complement fixation test for cattle vaccinated and infected with Brucella abortus.

A competitive enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody conjugated to horseradish peroxidase MA(A) and a complement fixation test (CFT) were applied to sera collected over a two-year period from 60 cattle challenged with Brucella abortus strain 544. Forty-eight of the cattle were previously vaccinated with B. abortus strain 19 (S19) or B. abortus strain 45/20 (45/20). After challenge 33 of the cattle remained uninfected and nine of the 27 infected cattle showed aberrant reactions by the CFT. The performance of the MA(A) ELISA was as follows: after vaccination, the MA(A) ELISA, like the CFT, was unable to differentiate infected cattle from those recently vaccinated with S19. After challenge the MA(A) ELISA gave results comparable with the CFT for those cattle with aberrant reactions. For the non-infected cattle there was a similar number of weeks after challenge when both tests were negative. It is suggested that the main advantage of the MA(A) ELISA when compared with the CFT lies in its relatively simple test procedure.